Genetic diversity of common carp from Shkodra Lake based on microsatellite markers

A Biba, A Hoda, E Vardhami and V Bozg

SHORT COMUNICATION

Abstract

Common carp (Cyprinus carpio) of Shkodra Lake is one of the most
important fish species. The genetic diversity of common carp in Shkodra Lake was
evaluated by the use of five microsatellite loci. A total of 30 individuals were
genotyped. The allelic number varied from 18 to 26. Mean observed heterozygosity
HO= 0.54 was lower than mean expected heterozygosity HE= 0.92. The population
was in Hardy Weinberg equilibrium for none of the loci. All the loci were highly
informative, mean PIC value was 0.926. The values of Shannon information index
(I) ranged from 2.5-3.3. The mean fixation index over the five loci was 0.421.
The bottleneck analyses showed no recent bottleneck.

Introduction

Shkodra Lake is the largest Lake in Balkan Peninsula, having a high variety
of fish species. There are about 60 fish species belonging to 17 families.
Cyprinids are one of the most important fish families based on the importance of
the species for the daily consume and the large number of species (Shumka et al
2008). Rich in plants and animals it represents an important ecosystem not only
for Albania but also at regional aspect. It is a reservoir of biodiversity and a migration route for many species in the region (Shumka et al 2013).

During the repopulation of the Lake there were used also fingerlings of other
invasive species (Flloko 2005, Shumka et al 2008), such as Carassius carassius
which competes for the habitat of common carp. This situation puts at risk the
sustainability of common carp populations (Dhora et al 2013; APAWA and CETI
2007). Hydrolisation and hybridization with improved races have changed the
morphometric characteristics of common carp in Shkodra Lake (Shermadhi and Spaho
2013). Another factor that put at risk carp population is overfishing and the
increase of illegal fishing during the spawning period (Dhora et al 2013; APAWA
and CETI 2007). During the period 2007-2011 the quantity of carp fished grew up
2-3 times normal values the demand for fish is increased with the growth of
population, economic level, processing industry, transport (Dhora et al 2013).

Carp of Shkodra Lake has been object of several studies. Golemi et al (2012,
2013) have studied biochemical and hematological parameters of this specie, and
the influence of sex in hematological and morphometric parameters of carp.
Evaluation of some metabolic and histopathological parameters were used to
detect the pollution stress of carp (Aliko et al 2012). Kolaneci et al (2009) has
studied the biological properties of carp population in Shkodra Lake. Shermadhi
and Spaho (2013) have studied the morphometric and meristic features variability
of carp of three Lakes. Berberi and Shumka (2013, 2014) studied clinical
indicators priory to diagnosis of spring viremia of carp and the detection of
spring viremia in common carp of Shkodra Lake.

In the current study, five microsatellite markers were used in order to
estimate the genetic diversity of Cyprinus carpio for Shkodra Lake.

Material and methods

Fish fins were taken from 30 individuals captured in the Albanian part of
Shkodra Lake and were used for the DNA isolation according to standard
procedures. Five microsatellite loci (MFW1, MFW6, MFW7, MFW18, MFW9) were
analyzed in these individuals. DNA fragments were amplified by polymerase chain
reactions (PCR) as described previously (Biba et al 2014). The amplified
fragments were separated by vertical electrophoresis in 6% denaturing
polyacrylamide gel using 4300 DNA Analyser (LICOR).

Average expected (HE) and observed (HO) heterozigosity values were calculated
by Genalex 6.5 program (Smouse and Peakall 2003). The same software was used to
evaluate Hardy Weinberg Equilibrium. Polymorphism information content (PIC)
values, which is a statistical assessment of informativeness of a marker, were
estimated by Cervus software (Marshall 1998). Factorial Correspondence Analysis
(FCA) was carried out by Genetix software v4.02 (Belkhir et al 2001). Bottleneck
software analyses were used to detect whether carp population of Shkodra Lake
has experienced a recent bottleneck, we used Wilcoxon sign-rank test (Piry et al
1999). Wilcoxon test has high power and can be used as few as four polymorphic
loci. Mode shift indicator, which discriminates bottlenecked populations from
stable populations, was used.

Results and Discussions

The allelic frequencies for the 5 loci are displayed in Figure 1.

Figure 1. Allelic frequencies
of five microsatellite loci.

Genetic diversity parameters are shown in Table 1. All markers were highly
polymorphic. PIC values for all markers were higher than 0.8, therefore these
markers appeared to be highly informative (Botstein et al 1980) and appropriate
in genetic diversity studies. A total of 69 alleles were observed at these 5
markers. All markers displayed significant departure from Hardy Weinberg
equilibrium, this may be due to non-random matting or natural selection. A
common cause of non-random matting is inbreeding. This is supported by
fixation index data. Observed heterozygosity (HO) ranged from 0.29
(MFW7) to 0.79 (MFW6) with a mean value of 0.54. Expected heterozygosity (HE)
ranged from 0.87 (MFW18) to 0.95 (MFW6) with a mean value of 0.92. High values
of expected heterozygosity reflect the presence of variation in this population.
Losses in heterozygosity values were published by Lehoczky et al 2005 (HO=0.557;
HE=0.700) using 12 microsatellite loci in six Hungarian strains of
common carp. Alam and Alam (2012) revealed also heterozygosity loss in three
captive strains of Bangladesh common carp analyzed by three microsatellite loci.
Fixation index values ranged from 0.17 (MFW6) to 0.69 (MFW7) with a mean value
of 0.42 indicating high level of heterozygote deficiency. This can be explained
by the presence of null alleles, Wahlund effect, and inbreeding. The values of
Shannon information index (I) ranged from 2.5-3.3.

Wilcoxon sign-rank test revealed probability values of 0.156
(IAM), 0.812 (TPM) and 0.625 (SMM) which were not significant (p>0.05), so null
hypothesis of mutation drift equilibrium was accepted under three models. These
results shows that Cyprinus carpio of Shkodra Lake did not experienced a recent
bottleneck. The same conclusion was also supported by the mode-shift indicator,
which showed a normal L-shape (Figure 2).

Factorial correspondence analysis (FCA) suggests that the first axis
accounted for 5.51%; second 5.26% and third 5.16% of the total variance (Figure
3) showing close genetic relationship between individuals. Only few individuals
show higher genetic distance and greater genetic diversity from the other part
of the population.

The whole population showed a high deficit of heterozygotes of 42.1%, which
supports also a close genetic relationship between sampled individuals displayed
by FCA.

Conclusion

All the markers used in the study were highly polymorphic.

None of the loci were in Hardy- Weinberg equilibrium.

All the 5 loci displayed losses in heterozygosity values.

No recent bottleneck was detected.

References

Alam N and Alam S 2012 Genetic varaition in three captive
populations of two strains of common carp (Cyprinus carpio L.) in Bangladesh
revealed by microsatellite DNA markers. Global Journal of Bio-Science and
Biotechnology 2: 223-228

Berberi E and Shumka S 2014 First detection of spring
viremia of carp virus in Cyprinus carpio, in Lake Shkodra/ Scadar Albania.
International Journal of Innovative Research in Science, Engineering and
Technology 3:18170-187175.